The present invention relates generally to DNA binding regulatory proteins and more particularly to DNA sequences encoding early growth regulatory proteins possessing histidine-cysteine "zinc finger" DNA binding domains, to the polypeptide products of recombinant expression of these DNA sequences, to peptides and polypeptides whose sequences are based on amino acid sequences deduced from these DNA sequences, to antibodies specific for such proteins and peptides, and to procedures for detection and quantification of such proteins and nucleic acids related thereto.
Among the most significant aspects of mammalian cell physiology yet to be elucidated is the precise manner in which growth factors (e.g., hormones, neurotransmitters and various developmental and differentiation factors) operate to effect the regulation of cell growth. The interaction of certain growth factors with surface receptors of resting cells appears to rapidly induce a cascade of biochemical events thought to result in nuclear activation of specific growth related genes, followed by ordered expression of other genes. Analysis of sequential activation and expression of genes during the transition from a resting state ("G.sub.0 ") to the initial growing state ("G.sub.1 ") has been the subject of substantial research. See, generally, Lau et al., Proc. Nat'l. Acad. Sci. (USA), 84, 1182-1186 (1987). Much of this research has involved analysis of the expression of known genes encoding suspected regulatory proteins (such as the proto-oncogenes, c-fos and c-myc) following mitogen stimulation. An alternative approach has involved attempts to identify genes activated by mitogenic stimuli through differential screening of cDNA libraries prepared from resting cells following exposure to serum and specific growth factors. See, e.g., Lau et al., EMBO Journal, 4, 3145-3151 (1985). See also, Cochran et. al., Cell, 33, 939-947 (1983), relating to the cloning of gene sequences apparently regulated by platelet derived growth factor.
Of interest to the background of the invention is the continuously expanding body of knowledge regarding structural components involved in the binding of regulatory proteins to DNA. Illustratively, the so-called receptor proteins are believed to bind to DNA by means of zinc ion stabilized secondary structural fingers premised on folding of continuous amino acid sequences showing high degrees of conservation of cysteines and histidines and hydrophobic residues. See, e.g., Gehring, TIBS, 12, 399-402 (1987). For example, a "zinc finger" domain or motif, present in Xenopus transcription factor IIIA (TF IIIA), as well as the Drosophila Kruppel gene product and various yeast proteins, involves "repeats" of about 30 amino acid residues wherein pairs of cysteine and histidine residues are coordinated around a central zinc ion and are thought to form finger-like structures which make contact with DNA. The histidine-cysteine (or "CC-HH") zinc finger motif, as opposed to a cysteine-cysteine ("CC-CC") motif of steroid receptors, is reducible to a consensus sequence represented as C-X.sub.2-4 -C-X.sub.3 -F-X.sub.5 -L-X.sub.2 -H-X.sub.3 -H wherein C represents cysteine, H represents histidine, F represents phenylalanine, L represents leucine and X represents any amino acid. [See, Klug et al. TIBS, 12, 464-469 (1987); Blumbeg et. al., Nature, 328, 443-445 (1987); and Schuh et al., Cell, 47, 1025-1032 (1986).]
Of particular interest to the background of the invention is the recent report of Chowdhury et al., Cell, 48, 771-778 (1987), relating to an asserted "family" of genes encoding proteins having histidine-cysteine finger structures. These genes, designated "mkrl" and "mkr2", appear to be the first such isolated from mammalian tissue and are not correlated to any early growth regulatory events.
There continues to exist a need in the art for information concerning the primary structural conformation of early growth regulatory proteins, especially DNA binding proteins, such as might be provided by knowledge of human and other mammalian DNA sequences encoding the same. Availability of such DNA sequences would make possible the application of recombinant methods to the large scale production of the proteins in procaryotic and eukaryotic host cells, as well as DNA-DNA and DNA-RNA hybridization procedures for the detection, quantification and/or isolation of nucleic acids associated with these and related proteins. Possession of such DNA-binding proteins and/or knowledge of the amino acid sequences of the same would allow, in turn, the development of monoclonal and polyclonal antibodies thereto (including antibodies to protein fragments or synthetic peptides modeled thereon) for use in immunological methods for the detection and quantification of early growth regulatory proteins in fluid and tissue samples as well as for tissue specific delivery of substances such as labels and therapeutic agents to cells expressing the proteins. In addition, DNA probes based on the DNA sequences for these mammalian early growth regulatory proteins may be of use in detecting gene markers used for the diagnosis of those clinical disorders which are linked to the marker genes.